Cockayne Syndrome (CS) is an autosomal recessive disorder, characterized by growth failure, neurological abnormalities, premature aging symptoms, and cutaneous photosensitivity, but no increased cancer incidence. CS is divided into two complementation groups: CSA (mutation in CKN1) and CSB (mutation in ERCC6). Of the patients suffering from CS, 80% have mutations in the CSB gene. We are pursuing the hypothesis that the primary role of CS proteins is to facilitate the repair of endogenous DNA damage, and we have evidence for a direct role of CSB in regulating BER efficiency. In addition, our recent work has revealed that Ca2+ is a novel metal cofactor for CSB catalyzed DNA-dependent ATP hydrolysis, but that CSB lacks detectable ATP dependent helicase and single- or double-stranded nucleic acid translocase activities in the presence of either Ca2+ or Mg2+. We have also discovered that (i) CSB supports ATP independent complementary strand annealing of not only DNA/DNA duplexes, but DNA/RNA and RNA/RNA duplexes;(ii) CSB forms a stable protein:DNA complex with a 34mer double-stranded DNA in electrophoretic mobility shift assays, independent of divalent metal or nucleotide (e.g. ATP);and (iii) CSB stably binds a range of nucleic acid substrates, including bubble and pseudo-triplex double-stranded DNAs that resemble replication and transcription intermediates, as well as forked duplexes of DNA/DNA, DNA/RNA, and RNA/RNA composition, the latter two of which do not promote CSB ATPase activity. Moreover, association of CSB with DNA, independent of ATP binding or hydrolysis, was found to displace or rearrange a stable pre-bound protein:DNA complex, a property likely important for its roles in transcription and DNA repair. More recent results, obtained in collaboration with Dr. Vilhelm Bohr, suggest that CSB plays a direct role in not only nuclear BER, but in mitochondrial BER, likely by helping recruit, stabilize, and/or retain BER proteins in repair complexes associated with the inner mitochondrial membrane. Future work will continue to examine the in vitro activities of CSB on key DNA and RNA transaction intermediates, and will elucidate the contributions of the unique N- and C-terminal portions of the protein that likely impart functional specificity. Huntington disease (HD) is a neurodegenerative disorder that belongs to a large family of genetic diseases caused by abnormal expansion of CAG/CTG repetitive sequences. Trinucleotide repeat expansions are unstable in the genome, both in germline and somatic cells. Expansion events in both cell types have deleterious clinical consequences in HD. For instance, transmission of longer repeats to offspring results in an earlier onset of disease, where extensive somatic expansion in the striatum, the brain region primarily affected in HD, is proposed to accelerate disease pathology. Thus, understanding the mechanisms of trinucleotide repeat instability is a major interest. We have recently found that oxidative DNA lesions abnormally accumulate at CAG expansions in a length-dependent, yet age- and tissue-independent, manner, likely due to the secondary structures formed by CAG repeats that limit access of enzymes that initiate BER. In addition, our data indicate that repair by BER enzymes of some of the accessible lesions results in somatic expansion when the ratio of FEN1 to POL is low, as found to occur in the striatum. Our results therefore support BER enzyme stoichiometry as a contributor to the tissue selectivity of somatic CAG expansion in HD, a hypothesis that we are currently pursuing in greater detail. XRCC1 is a critical scaffold protein that orchestrates efficient single-strand break repair (SSBR). Recent data has found an association of XRCC1 with proteins causally linked to human spinocerebellar ataxias - aprataxin and tyrosyl-DNA phosphodiesterase 1 - implicating SSBR in protection against neuronal cell loss and neurodegenerative disease. We have found that (i) shRNA lentiviral-mediated XRCC1 knockdown in human SH-SY5Y neuroblastoma cells results in a largely selective increase in sensitivity of the nondividing (i.e. terminally differentiated) cell population to the oxidizing agents, menadione and paraquat, and (ii) primary XRCC1 heterozygous mouse cerebellar granule cells or primary human fetal brain neurons depleted for XRCC1 exhibit increased strand break accumulation and reduced survival following menadione treatment. We are currently assessing the role of this protein in age-related pathologies using heterozygous mice, and will concomitantly evaluate the effect of XRCC1 haploinsufficiency on neurodegeneration and cancer proneness following defined insults. Finally, investigations exploring the possible role of aprataxin and tyrosyl-DNA phosphodiesterase 1 in maintaining mitochondrial function are ongoing.